Document Detail


Development of a quantitative assay for SARS coronavirus and correlation of GAPDH mRNA with SARS coronavirus in clinical specimens.
MedLine Citation:
PMID:  15735160     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
AIMS: To develop a quantitative reverse transcriptase polymerase chain reaction (Q-RT-PCR) for severe acute respiratory syndrome coronavirus (SARS-CoV) detection and explore the potential of using glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA as an internal control to exclude false negative results.
METHODS: SARS-CoV and GAPDH mRNA were both measured in 26 specimens from 16 patients with SARS, 40 follow up specimens from the same batch of patients, and appropriate control subjects. The relation between SARS positivity and GAPDH mRNA concentration was investigated using the chi2 test. Increasing the sensitivity for SARS-CoV and GAPDH mRNA detection was investigated in follow up specimens in which SARS-CoV and GAPDH mRNA were not detected initially.
RESULTS: Varying amounts of SARS-CoV were found in the 26 SARS-CoV positive specimens and SARS-CoV was not detected in the 40 follow up specimens and controls. In addition, concentrations of GAPDH mRNA were significantly different between the patients with SARS, follow up specimens, and healthy controls (Kruskal-Wallis test, p<0.05). Moreover, GAPDH mRNA concentrations were highly correlated with SARS-CoV positivity (chi2 = 5.43; p<0.05). Finally, SARS-CoV and GAPDH mRNA were both detected in three follow up urine specimens that were initially negative when the amount of cDNA used was increased from 5 microl to 10 and 15 microl.
CONCLUSIONS: This Q-RT-PCR assay can be used to detect SARS-CoV. Moreover, GAPDH mRNA may be useful to rule out false negative results in SARS-CoV detection, and the current extraction method for urine may not be sensitive enough to detect low titres of SARS-CoV.
Authors:
S C C Wong; J K C Chan; K C Lee; E S F Lo; D N C Tsang
Publication Detail:
Type:  Evaluation Studies; Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Journal of clinical pathology     Volume:  58     ISSN:  0021-9746     ISO Abbreviation:  J. Clin. Pathol.     Publication Date:  2005 Mar 
Date Detail:
Created Date:  2005-02-28     Completed Date:  2005-03-29     Revised Date:  2013-06-09    
Medline Journal Info:
Nlm Unique ID:  0376601     Medline TA:  J Clin Pathol     Country:  England    
Other Details:
Languages:  eng     Pagination:  276-80     Citation Subset:  AIM; IM    
Affiliation:
Department of Clinical Oncology, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong Special Administrative Region, China. cesar@clo.cuhk.edu.hk
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MeSH Terms
Descriptor/Qualifier:
Actins / biosynthesis,  genetics
Adult
Aged
Biological Markers / metabolism
False Negative Reactions
Female
Follow-Up Studies
Glyceraldehyde-3-Phosphate Dehydrogenases / biosynthesis*,  genetics
Humans
Male
Middle Aged
Quality Control
RNA, Messenger / genetics
RNA, Viral / genetics
Reverse Transcriptase Polymerase Chain Reaction / methods
SARS Virus / isolation & purification*
Sensitivity and Specificity
Severe Acute Respiratory Syndrome / diagnosis*
Chemical
Reg. No./Substance:
0/Actins; 0/Biological Markers; 0/RNA, Messenger; 0/RNA, Viral; EC 1.2.1.-/Glyceraldehyde-3-Phosphate Dehydrogenases
Comments/Corrections

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